טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentAxelman Elena
SubjectThe Effect of Vasopressin and Heparanase on Hepatocytes
Engraftment after Cell Transplantation in Rats
DepartmentDepartment of Medicine
Supervisor Clinical Professor Yaacov Baruch
Full Thesis textFull thesis text - English Version


Abstract

Background: Liver transplantation is considered the best treatment option for acute and chronic liver failure. However, shortage in the availability of donated liver organ has led researchers to develop alternative means. An attractive substitute is offered by transplantation of adult hepatocytes. This concept was evaluated under pre-clinical and clinical settings, yet its major limitation is low percentage (0.5-5%) of cell integration. Thus, increasing the effectiveness of hepatocytes transplantation is a major clinical and therapeutically challenge.

Aims: To study the role of heparanase in hepatocytes transplantation with or without prior partial hepatectomy (PHP). To study the effect of constitutively produced heparanase on short and long term hepatocyte transplantation. To examine the effect of vasopressin on hepatocytes transplantation.

Results: Over expression and secretion of heparanase in C6 cells were validated using enzymatic activity, PT-PCR, and immunoblot analyses. 24 hours following injection, transplanted hepatocytes were clearly identified in the spleen by H&E staining, and the number of spleen-residing hepatocytes was noted to be decline in subsequent days. This decrease was not affected by partial hepatectomy (PHP), or by the injection of hepatocytes and C6 cells, with and without heparanase. The results indicated a decrease in Y chromosome PCR product with time, in agreement with the morphological findings. Notably, we noted elevation of PCNA-positive cells in spleen tissues co-injected with hepatocytes and heparanase transfected C6 cells. Enhanced cell proliferation was in accordance with increased heparanase levels in the spleen tissue validated at the mRNA and protein levels.

Morphologically, transplanted hepatocytes were observed in portal areas of recipient livers. PCR analysis of Y chromosome revealed a marked increase in engrafted hepatocytes in recipient livers, an increase that again was restricted to rats subjected to PHP. The increase of native liver cells proliferation was restricted to rats co-injected with hepatocytes and heparanase transfected C6 cells, and only following PHP. Our results suggest that following PHP heparanase not only improves the engraftment of transplanted hepatocytes, but also the proliferation of native liver cells, contributing to the regeneration of injured liver.

Conclusions: Transplanted hepatocytes reside in the spleen for a relatively short period of time. Heparanase facilitates transplanted hepatocytes proliferation following partial hepatectomy in spleen. Efficient hepatocytes engraftment in the liver occurs only following partial hepatectomy and is induced by heparanase. Heparanase facilitates native liver cells proliferation following partial hepatectomy (hepatocytes and sinusoidal endothelial cells). Vasopressin did not improve hepatocytes engraftment in the liver in our animal model.